This invention relates to a process for the preparation of adenosine-5'-triphosphate by means of fermentation, and more particularly it relates to a process for preparing adenosine-5'-triphosphate according to a fermentation method by utilizing the microorganisms having methanol assimilability.
Growing interest is shown recently on availability of adenosine-5'-triphosphate (hereinafter referred to as ATP) for medicines, biochemical reagents, etc., as this substance plays an important role for the metabolism of energy as a high-energy phosphate compound in the living organism. Studies are being made for use of an enzyme pyruvate kinase, which is capable of regenerating adenosine-5'-diphosphate into ATP, in the ATP regeneration system for allowing efficient utilization of ATP in a bioreactor using ATP. Thus, uncostly supply of ATP is required for the production of the biochemical substances and coenzymes such as flavin adenine dinucleotide, nicotinamide adenine dinucelotide, etc.
There are known several methods for the production of ATP such as direct isolation from the animal muscle, organic chemical synthesis, enzymatic phosphatization of 5'-adenylic acid and fermentation. Regarding the last-mentioned fermentation method, there have been proposed some different techniques including a method in which a bacterium belonging to Brevibacterium ammoniagenes is cultivated in a medium containing adenine or a derivative thereof to thereby produce and accumulate ATP (Japanese Patent Publication No. 17634/1966), a method in which a microorganism having ATP producibility is cultivated in a medium containing a surfactant and ATP is collected from the cultures (Japanese Patent Publication No. 28996/1974), and a method in which a compound having a phenolic hydroxyl group is added in the medium at a point when the ATP yield in the medium has reached the maximum (Japanese Patent Laying-Open No. 6490/1978).
However, in any of these conventional methods adopting fermentation techniques, fermentation is effected by including adenine or a derivative thereof such as adenosine in the culture medium, that is, such adenine or a derivative thereof is used as substrate and it is phosphatized to thereby produce and accumulate ATP. Thus, the conventional methods use expensive adenine or its derivatives as starting material and hence can hardly be termed as industrially advantageous means.
As a result of further studies on means for producing ATP industrially at low cost by using the fermentation techniques, we found that ATP is accumulated in a high concentration in the culture medium when an ATP-producing bacterium having methanol assimilability is cultivated in a medium containing methanol or a chemical substance showing the same metabolic route as methanol as well as a specified amount of an inorganic phosphate as substrate instead of expensive adenine or its derivatives. This invention was reached on the basis of such finding.
The principal object of this invention, therefore, is to provide a process capable of producing ATP in an industrially advantageous way by means of fermentation.
Other objects of this invention will become apparent from the following detailed description of the invention.